INDUCTION OF KERATINOCYTE PROLIFERATION BY A SHORT TREATMENT WITH KERATINOCYTE-CONDITIONED MEDIUM

The effect of a short treatment with keratinocyte-conditioned medium ( KCM) on the growth of normal human epidermal keratinocytes was investi gated. Serum-free MCDB153 medium was conditioned by keratinocytes for 24 h after plating. Following attachment to uncoated plastic surfaces (4 h after plating), cells were exposed for 20 h to KCM. After 10 days of culture in MCDB153 medium, an increase of about six-fold in cell n umber was observed in KCM-treated plates over controls, indicating tha t a short treatment with KCM is sufficient to induce cell proliferatio n. The effect of addition of KCM at different times after plating was also evaluated: KCM treatment resulted to exert its maximum effect on cell proliferation, when performed immediately after the completion of attachment of cells to the surface of the dish. Mitogenic activity pr esent in KCM is not inhibited by heparin sulphate. The kinetics of acc umulation of this early secreted growth-stimulating activity showed th at a plateau is reached within 24 h of conditioning. These data sugges t that this mitogenic activity should not be amphiregulin. The observa tion that, following KCM treatment, the majority of cells is able to i ncorporate [H-3]-thymidine as compared to controls suggests that the o bserved final increase in cell number is due to an increase in the num ber of cycling cells rather than to a shortening of the cell cycle. (C ) 1997 Academic Press Limited.